Targeted vs. Untargeted Metabolomics

So I want to do metabolomics, now what?

Unfortunately, sample preparation and parameters for metabolomics experiments tend to be specific to your instrument and sample type, but there are some general considerations that you’ll need to consider before you begin. The first decision you’ll need to make is whether to perform targeted or untargeted metabolomics.

Targeted metabolomics looks at a specific number of known compounds (usually dozens to a couple hundred) in your sample. If done well, your samples will be run alongside pure standards for each of these known compounds to ensure that every molecule is correctly identified. If you already know which molecule(s) you are interested in, targeted metabolomics will provide the cleanest and most accurate results. But what if you don’t know ahead of time which metabolites are most important? 

That’s where untargeted metabolomics comes in. Instead of focusing on a handful of known compounds, untargeted metabolomics measures all the metabolites in your sample.* While this sounds great in theory, it can be quite complicated in practice. That’s because a single untargeted metabolomics run can measure tens of thousands of metabolites in a sample, many of which don’t match any known compounds in our databases. While there are a lot of cool ways to annotate these novel molecules (more on that later), untargeted metabolomics will leave you with a lot of unknowns. At Ometa Labs, we see these unknowns as opportunities for discovery, but we understand that if you just want to measure the concentration of methionine, untargeted metabolomics may provide a little too much information.

So how do I choose? It’s the classic conundrum of losing your keys on a dark road with a single streetlamp. Do you look for the keys where you think you dropped them, or do you look near the streetlamp because that’s where the light is? Targeted metabolomics is an excellent choice if you know what you’re looking for, but it will always lose information on all the metabolites you didn’t measure. Untargeted metabolomics can be harder to interpret, but keeps you from being limited to the small fraction of metabolites that have already been characterized.

TL;DR: if you already know exactly what metabolites you want to see, use targeted metabolomics. If you don’t or you’re unsure, use untargeted.

So how many metabolites are there anyways?

If you’re anything like me, the word metabolite brings back fond memories of trying to cram the citric acid cycle right before your biochemistry exam. But metabolites, and metabolism, go far beyond glycolysis. In recent decades, the advent of high-throughput, high-resolution mass spectrometry has vastly expanded our view of metabolism. In 2000, the recently-created KEGG pathway database contained 5,645 metabolites. Now, it contains over 19,000, while other repositories such as the human metabolome database (HMDB) contain more than 240,000. The Ometa Flow spectral library currently houses over 750,000 reference spectra and is constantly growing.

What these numbers don’t take into account is the fact that even these massive libraries often account for less than 15% of the molecules we can detect in a sample. This suggests that beyond the hundreds of thousands of molecules we already know, there are millions more that are still undiscovered. Starts to make your 200 metabolite panel look a little small, doesn’t it?

*For the pedants out there: no single method can measure every small molecule in a biological sample. Exactly which metabolites get measured can depend on things such as sample extraction, instrumentation, etc. But the idea behind untargeted metabolomics is to measure every metabolite you can see within the parameters of a particular experiment.

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